To meet the energy and food challenges of the 21st century, renewable and sustainable sources of fuel and nutrients are required. Microalgae offer promise in meeting these challenges but must achieve higher rates of areal productivity to compete with fully scaled, existing industries. In this study, we used selective O2 pressure to attain a ∼90 % increase in areal biomass productivity relative to the parental strain in the diatom Nitzschia inconspicua str. hildebrandi under pond-mimicking conditions with high O2 stress. The resulting culture line (GAI-337) was tested further for dilution time, culture density, CO2 supplementation, pH, temperature, and dissolved O2 concentration under outdoor pond-mimicking conditions to improve areal productivities. These experiments yielded an optimum harvest and dilution time just after sunset, ∼0.45 g AFDW L−1 initial culture density for maximal productivities, no requirement for CO2 gas supplementation or pH control, maximal performance under a diel temperature curve going from 24 °C at night to 36 °C during the day, and benefits from some O2 removal from the culture by bubbling with air. Using pond-mimicking laboratory bioreactors, N. inconspicua GAI-337 achieved ∼42 g AFDW m−2 d−1, placing it among the most productive microalgal strains tested to date. Nutrient limitation experiments resulted in a biomass composition with 36 % of AFDW comprised of lipids (measured as FAMEs) that equated to ∼160 Gallons of Gasoline Equivalent energy per ton AFDW, highlighting the potential of GAI337 as a promising renewable fuel feedstock strain.